In humans, gluten-sensitive enteropathy (GSE) is typically found in individuals genetically predisposed to celiac disease (CD), and in rhesus monkeys as well as in humans it can be induced by a gluten-containing diet. We recently performed experiments where gluten-sensitive and control macaques were fed gluten- containing diets followed by gluten-free diets. Complete recovery was achieved in GSE macaques - based on withdrawal of gluten from their diet. Furthermore, we identified 2 DRB haplotypes and/or 4 DQ allelic pairs as candidate MHC II genes for immunogenetic association with gluten sensitivity in rhesus macaques. Although several useful models have been established to study CD, including transgenic mice expressing HLA-DQ2 allele, there is no satisfactory animal model that would fulfill both genetic and pathologic criteria of this autoimmune disease. We have partially characterized a rhesus GSE model. We believe that such a model will be extremely useful for studies of the immunopathogenesis and treatment of CD. To develop this model further, we plan to: Aim 1 : Evaluate an association between clinical, histopathological and immunological surrogates of GSE in rhesus macaques. An association between clinical symptoms (diarrhea, weight loss, skin lesions, etc.), presence of AGA, anti-transglutaminase 2 (TG2) antibodies, villous atrophy, increased presence of IELs and inflammatory-cytokine producing intestinal T lymphocytes in macaques with clinical or subclinical GSE vs. controls will be evaluated. Aim 2 : Confirm MHC II alleles that were identified in rhesus macaques as candidates for immunogenetic association with gluten sensitivity. DNA extracted from at least 100 gluten-sensitive and 100 control macaques of Indian origin will be examined for the association with 2 DRB haplotypes and/or 4 DQ allelic pairs that we recently identified as MHC II candidate alleles. We predict that analogous to celiac patients DQ2/8 association will also be confirmed in rhesus macaques with GSE. Aim 3 : Evaluate the differences in a2-gliadin digestion between gluten sensitive and control macaques. In pilot study with gluten sensitive and control macaques, it was found that GSE animals but not controls, nor remitted animals, absorb undigested 33-mer across intestinal epithelium. Thus, it was proposed that systemic humoral immune response to dietary gluten is caused by absorption of undigested a2-gliadin across "leaky" epithelium in GSE macaques with proper MHC II type. Aim 4 : Evaluate the oral enzyme treatments in rhesus macaques with GSE. MHC II-pre- selected macaques with gluten sensitivity will be first placed on a gluten-free diet to accomplish remission. In a follow-up gluten challenge, gluten sensitive macaques will be dosed with increasing levels of gluten and a fixed daily oral dose of prolyl endopeptidase from Sphingomonas capsulata, cysteine endoprotease EP-B2 from barley, and the two-enzymes together to evaluate their therapeutic potential.

Public Health Relevance

It is estimated that more than one million Americans suffer from celiac sprue also known as celiac disease (CD). CD is an autoimmune disease of the small intestine caused by ingestion of gluten proteins from cereal grains. It was reported that there are clinical and immunological similarities between chronic inflammation of gastrointestinal tract in rhesus macaques (Macaca mulatta) and humans, suggesting the potential use of the non-human primates as a model for this NIDDK-relevant disease. The Gluten- Sensitive Enteropathy (GSE), accompanied with anti-gliadin antibodies (AGA), villous atrophy, reduced villus:crypt ratio and increased number of intraepithelial lymphocytes (IEL) can be identified in a relatively large proportion (~25%) of captive juvenile rhesus macaques of Indian origin with symptoms of idiopathic chronic diarrhea of non-infectious origin. In humans, GSE is typically found in individuals genetically predisposed to CD, and in rhesus monkeys as well as in humans it can be induced by a gluten-containing diet. We recently performed experiments where gluten-sensitive and control macaques were fed gluten-containing diets followed by gluten-free diets. It was concluded that complete remission could be achieved in GSE macaques - based on withdrawal of gluten from their diet. Furthermore, we identified 2 DRB haplotypes and/or 4 DQ allelic pairs as candidate MHC II genes for immunogenetic association with gluten sensitivity in rhesus macaques. The intestinal mucosa-associated lymphoid tissues can be sampled in non-human primates by non-invasive and repetitive endoscopic (pinch) biopsies to study specific questions with respect to pathogenesis and immunity. This cannot be accomplished in clinical studies with human patients or mice. Although several useful models have been established to study CD, including transgenic mice expressing HLA-DQ2 allele, there is no satisfactory animal model that would fulfill both genetic and pathologic criteria of this autoimmune disease. We have partially characterized a model of GSE in non-human primates. We believe that such a model will be extremely useful for studies of the immunopathogenesis and treatment of CD.